Each year, over 750,000 people in the United States suffer from severe sepsis and more than a third die. Sepsis is a phenotypic manifestation of infection and the host response, resulting in endothelial dysfunction and vascular injury. This contributes significantly to the development of multiple-organ failure and mortality. The mechanisms contributing to vascular injury in sepsis are multifaceted and extend beyond the influence of circulating factors elaborated from pathogens or the host's response with production of a myriad of cytokines and chemokines. Vascular and endothelial injury involves a complex interplay of innate and adaptive immune cells, platelets, coagulation factors, complement, and circulating plasma proteins. These cells and circulating molecules converge upon the endothelium to result in cellular activation and injury. This results in loss of an intact endothelium and endothelial homeostasis, vascular leakage, and inflammatory cell infiltration into parenchymal tissue. As a consequence, perfusion and oxygen delivery is altered and tissues are exposed to the same injurious factors, leading to parenchymal cell injury and dysfunction, and ultimately organ failure. The severe endothelial dysfunction that results in organ failure is now considered a hallmark of severe sepsis. Hemolysis in sepsis occurs to varying degrees and is not unique to specific pathogens. Red blood cell breakdown releases erythroid DAMPs (eDAMPs), including heme. These eDAMPs could have direct cytotoxic effects via oxidative injury or activate cell signaling pathways. Cell free hemoglobin and heme have been correlated with endothelial injury in a number of hemolysis-associated diseases. In sepsis, cell free hemoglobin correlates with outcomes in experimental models and is a predictor of survival in humans with severe sepsis. However, the role and mechanism(s) of eDAMPs in sepsis-induced endothelial injury remains unclear and is the focus of this proposal. Mitochondria have proven to be critical signaling organelles involved in a cells response to stress. eDAMPs and heme can influence mitochondrial signaling either directly or via PRRs. Furthermore, the influence of platelets and neutrophils as important amplifiers of endothelial injury will also be considered. These concepts will be considered in the context of the pulmonary endothelium and the development of acute lung injury, as lung injury arguably represents the most common clinical manifestations of organ injury in sepsis. Based on our published and preliminary data we propose the following hypothesis: Vascular cell and endothelial injury in sepsis is promoted by erythroid DAMPs, including heme. This hypothesis will be addressed by the following specific aims: Specific Aim 1: To determine the role and mechanism(s) of erythroid DAMPs to potentiate pulmonary endothelial injury in sepsis. Specific Aim 2: To determine the mitochondrial mechanism(s) by which eDAMPS induce endothelial injury and adaptation in sepsis. Specific Aim 3: To characterize mitochondrial function and dynamics in response to sepsis in human circulating cells and correlate plasma levels of eDAMPs with the inflammatory milieu, acute lung injury and outcomes in humans with severe intra-abdominal sepsis.